Compositions and Methods for Stimulation MAGP-1 to Improve the Appearance of Skin

ABSTRACT

The present disclosure relates to methods for treating, preventing and improving the condition and aesthetic appearance of skin, particularly, treating, preventing, ameliorating, reducing and/or eliminating fine lines and/or wrinkles of skin, by application of one or a combination of natural compounds or extracts that stimulate MAGP-1 expression and which improves the appearance of aged and/or photodamaged skin. The compositions of the invention are topically applied to the skin, or are delivered by directed means to a site in need thereof in an amount effective to improve the condition and aesthetic appearance of skin.

RELATED APPLICATIONS

This application claims priority to U.S. Provisional Patent ApplicationSer. No. 61/360,083, filed Jun. 30, 2010 and is a continuation of U.S.Nonprovisional patent application Ser. No. 13/158,947 filed on Jun. 13,2011, the contents of both of which are hereby incorporated by referencein their entirety.

FIELD OF INVENTION

The present invention relates generally to methods of improving theaesthetic appearance of human skin and/or improving the appearance ofaged and/or photodamaged skin by applying compounds to the skin whichstimulate the expression of the extracellular matrix protein MAGP-1 andprovide benefits to the skin.

BACKGROUND OF THE INVENTION

There is an increasing demand in the cosmetics industry to developproducts that may be applied topically to the skin that improve thecondition and appearance of skin. Consumers are interested in mitigatingor delaying the dermatological signs of chronologically- orhormonally-aged skin, as well as skin aging due to the environmentalstress, such as fine lines, wrinkles, sagging skin and other conditionsdue to a progressive loss of cell growth, proliferation andfunctionality in the epidermal and dermal skin layers. During the agingprocess, the complexion of the skin, i.e., the color and appearance ofthe skin, deteriorates slowly from aging and/or exposure toenvironmental stress, e.g., sunlight.

There remains a general need in the cosmetics industry for products thatretard or counter the aging effects on the skin, and more specificallyfor products that produce such effects without undesirable side effects.In particular, there remains a need for topically applied cosmeticcompositions that provide anti-aging and skin texture benefits usingnatural plant materials as active components.

Active ingredients or components derived or extracted from plants andplant seeds have commonly been employed for a myriad of medicinal,therapeutic and cosmetic purposes. Such actives may be obtained from theentire plant or various parts of a plant, such as seeds, needles,leaves, flowers, roots, bark, cones, stems, rhizomes, callus cells,protoplasts, and meristems. Active ingredients or components areincorporated in compositions in a variety of forms. Such forms include apure or semi-pure component, a solid or liquid extract or derivative, ora solid natural plant material. Plant material may be incorporated in avariety of subforms such as whole, minced, ground or crushed, orotherwise physically modified for incorporation into a composition.

MAGP-1 (Microfibril-associated glycoprotein 1) is a small glycine richacidic protein in extra-cellular matrix (ECM). MAGP-1 is a 30-33 kDamember of the microfibril associated glycoprotein family of proteins. Itwas found to be a key component of microfibrils and elastic fibers inskin. MAGP-1 forms complexes with other microfibril proteins such asfibrillin. Besides providing structural support for skin elastic fiberformation, MAGP-1 is also essential for vascular integrity, woundhealing, and proper body fat deposition, possibly through regulatingTGF-beta signaling, based on animal studies. MAGP-1 apparently mediatesthe release of Notch extracellular domain and also likely binds to typeIV collagen and contributes to elastin fiber formation.

The present invention identifies a number of compounds and/or plantmaterials and extracts found to be active at simulating expression ofthe extracellular matrix protein MAGP-1. Among the plant materials,which preferably are in the form of extracts, are Sesbania grandiflora,Antidesma bunius, Operculina turpethum, Ixora chinensis, Clerodendronlindleyi, Amorphophallus campanulatus, Tiliacora triandra, andThunbergia laurifolia. Suitable compounds are the tetrapeptidelysine-threonine-phenylalanine-lysine (Lys-Thr-Phe-Lys) (herein KTFK),thiodipropionic acid (TDPA) and its lower alkyl mono- and diesters, asubstituted 2-(2-benzyl-1-benzimidazoyl) acetamide analog having thestructure:

and an N-substituted sulfonyloxybenzylamine having the structure:

Certain of the MAGP-1 active plant species, namely, Antidesma bunius,Operculina turpethum, Ixora chinensis, and Clerodendron linleyi,including extracts of these plant species, have not been previouslyknown for topical application to skin to improve the skin's aestheticappearance. Similarly, the above-disclosed substituted2-(2-benzyl-1-benzimidazoyl) acetamide analog and the N-substitutedsulfonyloxybenzylamine as disclosed above have not been known previouslyfor topical application to skin to improve the skin's aestheticappearance.

Antidesma bunius is a species of fruit tree in the spurge family. It isnative to Southeast Asia, the Philippines, and northern Australia. Itscommon Philippine name and other names include bignay, bugnay or bignaiand currant tree. This is a variable plant which may be short andshrubby or tall and erect, approaching 30 meters in height. It has largeoval shaped leathery evergreen up to about 20 centimeters long and sevenwide. The leaves are sudorific and employed in treating snakebite, inAsia. (Morton, J. 1987. Bignay. p. 210-212. In: Fruits of warm climates.Julia F. Morton, Miami, Fla.).

Operculina turpethum is a plant in the morning glory family, (syn.Ipomoea turpethum) and is known commonly as turpeth, fue vao, and St.Thomas lidpod. It is one of many plants mentioned in the literaturehaving claims of activity against liver disorders. Vasudesan, N, V.,Indian Medicinal Plants, Vol. IV, Orient Longman Ltd, Chennai, 1995,172. It also has anthelmintic expectorant, antipyretic,anti-inflammatory and purgative properties. Id.

Ixora chinensis is a fairly small, bushy shrub, usually only 5-10 fttall. There are dwarf varieties that are much smaller. Ixora will flowerwhen only a few inches high. It is commonly used a hedge or small gardenplant. The plant is mainly grown as ornamental but its flowers arebelieved to have some medicinal uses including treatment forhypertension and for treating rheumatism, abscesses, bruises and wounds.The plant is native to India and Sri Lanka

Clerodendron lindleyi are shrubs, lianas, and small trees, usuallygrowing to 1-12 m tall, with opposite or whorled leaves. The genus isnative to tropical and warm temperate regions of the world, with most ofthe species occurring in tropical Africa and southern Asia, but with afew in the tropical Americas and northern Australasia, and a fewextending north into the temperate zone in eastern Asia. Clerodendrum isfrom the Greek, meaning “lottery tree”. Dendros meaning tree. Thelottery refers to unsure possibility of medicinal value from certainplants of this genus.

Sesbania grandiflora (also known as agati, syn. Aeschynomenegrandiflora) or hummingbird tree/scarlet wisteria is a small tree in thegenus Sesbania. It is a fast-growing tree with a typical adult height ofbetween 3 and 5 m. The leaves are regular and rounded and the flowerswhite and large, very characteristic. The fruits look like flat, longand thin green beans. The tree thrives under full exposure to sunshineand is extremely frost sensitive. It is believed to have originatedeither in India or Southeast Asia and grows primarily in hot and humidtropical areas of the world. Bark, leaves, gums, and flowers areconsidered medicinal. The astringent bark was used in treating smallpoxand other eruptive fevers. (James A. Duke. 1983. Handbook of EnergyCrops. unpublished.) Sesbania grandiflora was previously disclosed inU.S. Pat. No. 7,618,662, the disclosure of which is incorporated hereinby reference.

Thunbergia laurifolia or Blue trumpet vine is an invasive species, whichis used like an ornamental plant. This plant is found in Cerradovegetation of Brazil. In Malaysia, juice from crushed leaves of T.laurifolia are taken for menorrhagia, placed into the ear for deafness,and applied for poulticing cuts and boils (Burkill, I. H. (1966). “ADictionary of the Economic Products of the Malay Peninsula. Volume II(I-Z)”. Ministry of Agriculture and Cooperatives, Kuala Lumpur). InThailand, leaves are used as an antipyretic, as well as an antidote fordetoxifying poisons (Kanchanapoom, T. et al. (2002). “Iridoid glucosidesfrom Thunbergia laurifolia”. Phytochemistry 60: 769-771).

KTFK is a synthetic tetrapeptide lysine-threonine-phenylalanine-lysine(Lys-Thr-Phe-Lys), known to be active in collagen and fibronectinsynthesis. KTFK was previously disclosed in U.S. Patent ApplicationPublication No. 2004/0126344, the disclosure of which is incorporatedherein by reference.

Amorphophallus campanulatus is found in tropical regions from Madagascarto Asia, Polynesia and northern Australia. The root is carminative,restorative, stomachic and tonic. Chopra. R. N., Nayar. S. L. andChopra. I. C. Glossary of Indian Medicinal Plants (Including theSupplement). Council of Scientific and Industrial Research, New Delhi.1986; Medicinal Plants of Nepal Dept. of Medicinal Plants. Nepal. 1993.It is dried and used in the treatment of piles and dysentery. Id. Thefresh root acts as an acrid stimulant and expectorant, it is much usedin India in the treatment of acute rheumatism. Id. Amorphalluscampanulatus was previously disclosed in U.S. Pat. No. 7,618,862, thedisclosure of which is incorporated herein by reference.

Thiodopropionic acid (TDPA) is white crystalline solid having a slightcharacteristic odor and has demonstrated some antimicrobial andantioxidants activity. (WHO/Food Add./24.65 FAO Nutrition MeetingsReport Series No. 38A). Thiodopropionic acid and its esters waspreviously disclosed in U.S. Patent Application Publication No.2004/0126344, the disclosure of which is incorporated herein byreference.

Sapindus rarak is part of the Sapindus genus comprising about five totwelve species of shrubs and small trees in the Lychee family,Sapindaceae, native to warm temperate to tropical regions in both theOld World and New World. The extract of Sapindus rarak has demonstratedanalgesic effects. Sapindus rarak was previously disclosed in U.S. Pat.No. 7,514,092, the disclosure of which is incorporated herein byreference.

Tiliacora triandra Diels of the Tiliacora family, also known as Yanang,is a species of flowering plant native to mainland Southeast Asia andused particularly in the cuisines of northeast Thailand and Laos. It isa climbing plant with mostly single, smooth, oval-shaped, deep greenleaves and yellowish flowers. In traditional Southeast Asian medicine,Tiliacora triandra has been used as an herbal medicine for fever relief,alcohol intoxication, inflammation, and bacterial/fungal infection. Forinstance, the use of Tiliacora triandra Diels against plasmodiumfalciparum (cause malaria in humans) is disclosed in Pavanand et al.,Phytother. Res., 3, 215-217 (1989). Tiliacora triandra is the subject ofa separate application filed concurrently with the instant application.

Safe, effective and new components of compositions to treat, prevent,reduce, inhibit and/or improve the dermatological signs of aging,including environmental stress, due to a progressive degradation of theepidermal and dermal skin layers, would be advantageous for theformulation of treatments and products for the skin. As describedherein, novel and beneficial methods and compositions, as well as theirmode of action, for the treatment of wrinkles and the like, as well asfor personal care products for the skin, are provided herein.

SUMMARY OF THE INVENTION

It is an object of the present disclosure to provide a topicalcomposition having a compound, particularly a natural plant material,blends thereof, or components therefrom, in an amount sufficient toimprove the aesthetic appearance of human skin, and to prevent,ameliorate and/or reduce dermatological signs of chronologically orhormonally-aged or photo-aged skin, such as fine lines, wrinkles,sagging skin, and other conditions due to a progressive degradation ofthe skin cell growth, proliferation and functionality in the epidermaland dermal layer, where the plant material, blends thereof or componentstherefrom are in a cosmetically, dermatologically acceptable vehicle,carrier, or diluent.

It is another object of the present invention to provide compositionssuitable for topical application to human skin having one or more MAGP-1active agents that stimulate the expression of MAGP-1 comprising: (i) aplant material, especially extracts thereof, selected from Antidesmabunius, Operculina turpethum, Ixora chinensis, and Clerodendronlindleyi, and mixtures thereof, and (ii) a compound selected from2-(2-benzyl-1-benzimidazoyl) acetamide analogs and an N-substitutedsulfonyloxybenzylamine, and combinations/mixtures thereof.

It is another object of the present invention to provide a method oftopically applying to human skin a composition having a compound,particularly a natural plant material, blends thereof, or componentstherefrom and which stimulate the expression of MAGP-1.

It is further object of the present disclosure to provide topicalcompositions comprising two or more ingredients wherein at least onefirst ingredient is selected from the group consisting of Antidesmabunius, Operculina turpethum Ixora chinensis, Clerodendron lindleyi, andan N-substituted sulfonyloxybenzylamine, and at least one secondingredient is selected from the group consisting of Sesbaniagrandiflora, Amorphophallus campanulatus, Sapindus rarak, Tiliacoratriandra, Thunbergia laurifolia, tetrapeptidelysine-threonine-phenylalanine-lysine (KTFK), and Thiodipropionic acid(TDPA) and its esters, in an amount effective to treat, prevent,control, ameliorate, inhibit, and/or reduce dermatological signs ofchronologically or hormonally-aged or photo-aged skin, such as finelines, wrinkles, sagging skin, and other conditions due to a progressivedegradation of the skin cell growth, proliferation and functionality inthe epidermal and dermal layer.

It is still a further object of the present disclosure to provide amethod of improving the aesthetic appearance of skin, including treatingthe effects of aging in the skin, by topically applying the compositionsof the disclosure to the skin.

It is an additional object of the present disclosure to provide ascreening method for identifying compounds that stimulate the expressionof the MAGP-1 protein and to provide compositions using the activesidentified in such screening method.

Further objects, features and advantages of the present disclosure willbe better appreciated upon a reading of the detailed description.

In accordance with the foregoing objectives and others, it hassurprisingly been found that expression of MAGP-1 in skin improves theappearance of aged or photodamaged skin by increasing elasticity,tightening pores, improving blood vessel integrity and reducing undereyedark circle, and reducing overall lines, wrinkles, and sagging.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 illustrates the MAGP-1 protein expression in dermis ofphoto-protected and photo-exposed younger subjects (age 18-25) and oldersubjects (age 40-65).

FIG. 2 illustrates the MAGP-1 expression around hair follicles and poresof the photo-protected and photo-exposed younger subjects (age 18-25)and older subjects (age 40-65).

DETAILED DESCRIPTION

All terms used herein are intended to have their ordinary meaning unlessotherwise provided.

The present disclosure provides novel active agents and novel methods ofimproving the aesthetic appearance of human skin by topically applyingcompositions newly found to be effective to treat signs of aging andresults relating to the dermatological aging of skin, such as finelines, wrinkles, sagging skin, and other conditions, due to aging,including chronological aging, hormonal aging and/or photo-aging, toimprove the aesthetic appearance of skin. It is to be understood thatchronological aging represents the structural, functional, and metabolicchanges in the skin that parallel the aging and degenerative change inother body organs, whereas photo-aging is a separate process and largelyinvolves damage to the collagen and elastin fibers in the skin due to anexposure to environment such as the sunlight. Improvements in theaesthetic appearance of the skin may be achieved by topical applicationof these compositions to the skin on a regular and consistent basis,such as daily basis.

It has been found, as demonstrated in FIGS. 1 and 2, and described ingreater detail in the Examples, that MAGP-1 in skin tissue changes withage and photoexposure. Accordingly, aging skin will manifest a reductionin the number of MAGP-1 complexes formed with fibrillin and othermicrofibril proteins, with concomitant loss of skin elasticity, andnumerous other properties of healthy skin. The compositions of thepresent invention containing one or more of the active agents forincreasing MAGP-1 expression are provided to reduce, lessen, amelioratethe harmful effects of aging of skin, and to help restore a moreyouthful appearance to the skin.

Without wishing to be bound by theory, compounds and natural plantmaterials and extracts described herein (hereinafter, the MAGP-1 activeagents) stimulate the expression of MAGP-1 in skin and improve theappearance of aged or photodamaged skin by increasing elasticity,tightening pores, improving blood vessel integrity, reducing undereyedark circles and reducing overall lines, wrinkles and skin sagging. Itis believed that the expression of MAGP-1 forms complexes withmicrofibril proteins such as fibrillin. Fibrillin is a glycoprotein,which is essential for the formation of elastic fibers found inconnective tissue. Fibrillin is secreted into the extracellular matrixby fibroblasts and becomes incorporated into the insoluble microfibrils,which appear to provide a scaffold for deposition of elastin the proteinin connective tissue that is elastic and allows many tissues in the bodyto resume their shape after stretching or contracting.

The present disclosure provides novel compositions and their methods ofuse newly found to be effective to treat signs of aging and resultsrelating to the dermatological aging including chronological aging,hormonal aging and/or photo-aging, of skin by effecting one or more ofincreasing elasticity, reducing fine lines, wrinkles, sagging skin,reducing undereye darkness/bruising and fat pockets, trimming/slimmingthe effect of the fat pad improving skin tone, providing improvedradiance and healthy glow, and other conditions.

Inasmuch as MAGP-1 stimulation increases formation of microfibrilproteins such as fibrillin, which is secreted into the extraacellularmatrix by fibroblasts, it is a further embodiment of the invention toprovide compositions that contain the MAGP-1 active agents of thepresent invention in combination with materials, plant or compounds,that have activity to increase proliferation of fibroblasts. Suchcombination increases production of collagen; provides an increase inanchoring and adhesion between keratinocytes, promotes cell-celladhesion in the epidermis and dermis, and strengthens dermal-epidermaljunctions. Collaterally, there is an improvement in the aestheticappearance of the aging skin evidenced by an increase in one or more ofskin firmness, skin plumpness, skin suppleness, or skin thickness.

The present invention further provides methods for topically applying acomposition to the skin which improve the condition and appearance ofskin by stimulating the expression of the extracellular protein MAGP-1.

The methods in accordance with the present invention incorporate one ora combination of MAGP-1 active agents comprising the compounds and/orplant extracts from the group consisting of Sesbania grandiflora,Antidesma bunius, Operculina turpethum, Ixora chinensis, Clerodendronlindleyi, tetrapeptide lysine-threonine-phenylalanine-lysine (KTFK),Amorphophallus campanulatus, thiodipropionic acid (TDPA) and its mono-and diesters of from 1 to 6 carbon atoms, Sapindus rarak, Tiliacoratriandra, Thunbergia laurifolia, the N-substitutedsulfonyloxybenzylamine having the structural formula I, namely,4-((N-isobutylphenylsulfonamido)methyl)phenyl methanesulfonate (CAS853758-16-4):

and the substituted 2-(2-benzyl-1-benzimidazoyl) acetamide analog havingthe structural formula II, namely,2-{2-[(3-methylphenyl)methyl]-1H-1,3-benzodiazol-1-yl}1-(morpholin-4-yl)ethan-1-one(CAS 1043743-29-8):

Compounds I and II are commercially available from AMRI Hungary, Zahonyu 7, H-1031 Budapest, Hungary

It is to be understood that, as used herein, the terms treating andtreatment include and encompass reducing, ameliorating, improving,alleviating, and/or eliminating the dermatological effects of agingand/or environmental stress. The present compositions and methods aresuitable for use in treating cosmetic and dermatological conditions ofthe skin in numerous areas of the body, including, without limitation,the face, forehead, lips, neck, arms, hands, legs, knees, feet, chest,back, groin, buttocks, and the like. In a preferred embodiment, thecompositions are applied to the face.

One embodiment of the present disclosure relates to novel compositionscomprising one or a combination of novel MAGP-1 active agents selectedfrom the group of Antidesma bunius, the N-substitutedsulfonyloxybenzylamine of structure I (above), the substituted2-(2-benzyl-1-benzimidazoyl) acetamide analog of structure II (above),Operculina turpethum, Ixora chinensis, and Clerodendron lindleyi, and acosmetically, dermatologically, or pharmacologically acceptable vehicle,and to methods for improving skin by topically applying to skin in needof treatment said novel topical compositions.

In another embodiment compositions of the present invention comprise oneor more of the novel MAGP-1 active agents set forth above in combinationwith one or more other MAGP-1 active agents, and a cosmetically,dermatologically, or pharmacologically acceptable vehicle, and tomethods for improving skin by topically applying to skin in need oftreatment said novel topical compositions. Among the one or more otherMAGP-1 active agents mention may be made of Sesbania grandiflora, KTFK,Amorphophallus campanulatus, thiodipropionic acid (TDPA) and its C₁ toC₆ alkyl mono- and diesters, Tiliacora triandra, and Thunbergialaurifolia.

Histological studies of the skin show that as aging occurs, the skinundergoes structural, functional, and metabolic changes that parallelthe aging and degenerative changes in other body organs. Whilechronological and/or hormonal aging play a significant role, the numberof wrinkles present is also highly dependent on the amount of exposureto environmental stress during an individual's life, e.g., repetitivesun exposure over 10, 20, 30, or more years of the person's lifeoccasions oxidative damage from overexposure to ultraviolet (UV)sunlight. While there is a gradual thinning of male skin with increasingage of approximately 1% per year, the thickness of most women's skinsremains surprisingly constant until the menopause, after which there isa significant and sometimes dramatic thinning with increasing age,reinforcing the need in older women for a treatment that increases thecollagen content of skin and hence improves the firmness, thickness andplumpness of skin.

Thus, the successful restoration of youthful skin from this perspectivemust address a variety of key issues including: vitality of fibroblastsand keratinocytes, cell-cell adhesion in the epidermis and dermis, cellnourishment to the epidermis, cell-cell anchoring and adhesion betweenkeratinocytes, communication between the dermis and epidermis,collagenase overproduction, collagen replacement, and mechanicalproperties of the skin. Cosmetic ingredients that addresses these keyissues, are useful in the topical composition of the present disclosure.

Topical Compositions

The natural plant material may be in any form including, but not limitedto, the whole plant, a dried plant, a ground plant, or parts thereof,including but not limited to, seeds, needles, leaves, roots, bark,cones, stems, rhizomes, callus cells, protoplasts, flowers, andmeristems, or components and/or constituents found in, or isolated from,the natural plant material, and/or portions of the plant, or anycombinations thereof. In one embodiment, the natural plant material isin the form of an extract derived from a select portion of the naturalplant material. Typically, the plant extract is obtained from the wholeplant or form the leaves of the plant. It is to be understood that“natural plant material” also includes an ingredient, component,constituent, or extract derived from the natural plant material.

In the case of Tiliacora triandra, the plant extract is preferablyobtained from the vines of the plant. In the case of Ixora chinensis andSesbania grandiflora, the extract is preferably obtained from theflowers of the plant. In the case of Antidesma bunius, Thunbergialaurifolia, and Clerodendron lindleyi the extract is preferably obtainedfrom leaves. In the case of Operculina turpethum the extract ispreferably obtained from the aerial portion of the plant, i.e., thewhole plant excluding the roots.

Insofar as Tiliacora triandra compositions are concerned in connectionwith the present invention, the present invention contemplatesembodiments excluding hydrogen peroxide, iodine, tarrow root and amixture of tarrow root and iodine.

The composition comprising these plants is effective when topicallyapplied, preferably in a daily manner.

The compositions have a concentration of the MAGP-1 active agents offrom about 0.0001 wt % to about 90 wt %, about 0.001 wt % to about 25 wt%, about 0.01 wt % to about 10 wt % based on the total weight of thecomposition, about 0.05 wt % to about 1 wt %. In the case of plantextracts, the above amounts refer to an “active amount” of the plantextract. The terms “active amount” or “dry weight” are used synonymouslyand refer to the amount of the plant extract after solvent and/or otherdiluents have been removed. One of ordinary skill in the art would beable to adjust the amount of extract used based upon the specificapplication or effect desired.

The MAGP-1 active agents of the present disclosure are preferablycontained in a cosmetically or dermatologically acceptable vehicle,medium, diluent or carrier. In an embodiment embracing topicalapplication, the compositions of this disclosure comprise a medium(vehicle, diluent or carrier) that is compatible with human skin. Thecompositions can be formulated as aqueous, alcohol, oraqueous/alcohol-based solutions, ointments, lotions, gels, water-in-oil,oil-in-water, of water-oil-water triple emulsions having the appearanceof a cream or gel, microemulsions, or aerosols. In addition, thecompositions can be in the form of vesicular dispersions containingionic and/or nonionic lipids, as described above. Dosage units suitablefor such compositions are formulated according to the conventionalknowledge and techniques used in the art.

Methods of Using MAGP-1 Active Agents

As will be appreciated by the practitioner, cosmetic treatmentscomprising compositions containing natural plant materials, includingextracts, components, and/or constituents of the invention may becarried out, for example, by topically applying the cosmetic compositionas described herein according to a regimen for administering suchcompositions. The topical cosmetic composition preferably is appliedonce or twice daily (e.g., morning and evening) for a period of at leastone week, but may include a period of about 2, 4, 8, or 12 weeks. Theconsumer may wish to continue use of the composition for an extendedperiod of time. The cosmetic composition is preferably applied to theface and neck, but may be applied to any area of skin in need ofaesthetic improvement, where the cosmetic composition remains on theaffected area of skin, and preferably not removed or rinsed off theskin. The methods encompass the application of compositions/products inthe form of creams, lotions, gels, masks, sera, ointments, patches,makeup, makeup-removing milks, sunscreen compositions, or the like, tothe skin. Preferably the cosmetic composition is a topical leave onformulation, where spraying as a form of application is also envisioned.

In a particular embodiment of the invention, the topical compositionshaving an MAGP-1 active agent of the present invention, includingcompositions containing two or more of such agents, and compositionscontaining one or more novel active agents as previously defined, aloneor in combination with one or more other active agents, includingcomponents or extracts derived therefrom, are useful for improving thecondition and aesthetic appearance of skin affected by aging,particularly matured or maturing skin, by anyone of the followingmethods: reducing dermatological signs of chronological aging,photo-aging, hormonal aging, and/or actinic aging; preventing and/orreducing the appearance of lines and/or wrinkles; reducing thenoticeability of facial lines and wrinkles, facial wrinkles on thecheeks, forehead, perpendicular wrinkles between the eyes, horizontalwrinkles above the eyes, and around the mouth, marionette lines, andparticularly deep wrinkles or creases; preventing, reducing, and/ordiminishing the appearance and/or depth of lines and/or wrinkles;improving the appearance of suborbital lines and/or periorbital lines;reducing the appearance of crow's feet; rejuvenating and/or revitalizingskin, particularly aging skin; reducing skin fragility; ameliorating theeffects of estrogen imbalance; preventing and/or treating skin atrophy;preventing, reducing, and/or treating hyperpigmentation; minimizing skindiscoloration; improving skin tone, radiance, clarity and/or tautness;preventing, reducing, and/or ameliorating skin sagging; improving skinfirmness, plumpness, suppleness and/or softness; improving procollagenand/or collagen production; improving skin texture and/or promotingretexturization; improving skin barrier repair and/or function;improving the appearance of skin contours; restoring skin luster and/orbrightness; minimizing dermatological signs of fatigue and/or stress;resisting environmental stress; replenishing ingredients in the skindecreased by aging and/or menopause; improving communication among skincells; increasing cell proliferation and/or multiplication; increasingskin cell metabolism decreased by aging and/or menopause; retardingcellular aging; improving skin moisturization; enhancing skin thickness;increasing skin elasticity and/or resiliency; enhancing exfoliation;improving microcirculation; decreasing and/or preventing celluliteformation; and any combinations thereof.

The active components of the disclosed topical composition are furtheruseful in treating, preventing, arresting, ameliorating, reducing ordiminishing medical and/or dermatological conditions of skin affected byaging. Such conditions, as used herein, commonly include, but are notlimited to, dermatological aging (chronological aging, hormonal agingand/or actinic aging), dermatitis, skin and hair fragility, rosacea,skin blemishes, sensitive skin, hyperpigmentation or hypopigmentation,thinning skin, roughness, keratosis, skin atrophy, wrinkles, lines,hyperplasia, fibrosis, and any combinations thereof. The activecomponents of the topical composition may also be useful in enhancingthe general health, vitality, condition, and appearance of the skin.

Gene expression may be measured by the determination of RNA levels incultured cells, for example, using techniques such as Northern blottechnology and the polymerase chain reaction (PCR), e.g., “real time”PCR and reverse transcription PCR (RT PCR) as practiced in the art.(see, e.g., Sambrook et al., 1989, Molecular Cloning: A LaboratoryManual, Cold Spring Harbor Laboratory, Cold Spring Harbor, New York; R.Higuchi et al., 1992, Biotechnology, 10:413-417; R. Higuchi et al.,1993, Biotechnology, 11: 1026-1030; E. S. Kawasaki, 1990, “Amplificationof RNA”, In: RNA Protocols: A Guide to Methods & Applications, M. A.Innis et al., Academic Press, San Diego, Calif., pp. 21-27; all of whichare herein incorporated by reference). In addition, gene expression inskin, skin substitute, or cultured cells may be evaluated using gene(cDNA) arrays (microarrays or nucleic acid gene chip test arrayscomprising membrane, glass, plastic or silicon support materials or thelike), serial analysis of gene expression (SAGE), (e.g., as described byV. E. Velculescu et al., Science, 270(5235):484-487, 1995; A. Lal etal., Cancer Res., 59(21):5403-5407, 1999; both of which are hereinincorporated by reference), or differential display techniques all ofwhich are commonly known and used in the art.

The topical compositions use plant materials/active ingredients affectedby genes associated with dermatological signs of aging, such as finelines, wrinkles, and sagging skin as a biomarker for compounds which mayimprove the condition and appearance of affected skin. If expressionlevels of such nucleic acid/protein biomarkers are modified or elevatedin the presence of a natural plant material or active ingredienttherefrom, the natural plant material may be used in a topicalcomposition of the invention for improving the condition and appearanceof skin. Such expression level assays embrace a variety of methods formeasuring nucleic acid levels in cells that have been exposed to one ormore test substances. Suitable methods include detection and evaluationof gene activation or expression of, for example, DNA, RNA, or mRNA. Asnon limiting examples, polymerase chain reaction (PCR) assays (e.g., RT-PCR), Northern blotting, in situ hybridization, and other assays asknown and practiced in the art may be employed to quantify RNA in cellsbeing assayed for tolerance to a particular treatment (see, e.g., J.O′Connell, 2002, RT-PCR Protocols, Humana Press, Totowa, N.J.; R. Rapleyand D. L. Manning, 1998, RNA Isolation and Characterization Protocols,Humana Press; R. Rapley, 2000, Nucleic Acid Protocols Handbook, HumanaPress; all of which are herein incorporated by reference). In accordancewith such assays, if levels of at least one nucleic acid biomarker areelevated in the presence of one or more test substances, this maypredict that the substance(s) will improve the dermatological signs ofaging. These substances, or natural plant materials, may then be used ina topical composition, preferably applied daily to the skin, in order totreat, prevent, ameliorate, and/or reduce, signs of dermatologicalaging, especially fine lines, wrinkles, and sagging skin, therebyimproving the condition and aesthetic appearance of skin.

In another embodiment, the plant extract as used herein, also includes“synthetic” extracts, i.e., various combinations of known plantcomponents and/or constituents that are combined to substantially mimicthe composition and/or activity of a plant extract of natural origin.Such synthetic extracts are included in the term “plant extract”. Thesynthetic extracts will have two or more, three or more, or four or moreactive ingredients in common with a plant. Most preferably, thesynthetic extracts will have substantially the same number of activeingredients as a natural extract. The correspondence of the numericalincidence of active ingredients between the synthetic extracts and theplant or a natural extract may also be described in terms of “percentcommonality”. Preferably, the synthetic extract has about 50 percent ormore commonality to the chemical composition of a plant or naturalextract. In other words, the synthetic extract has about 50 percent ormore of the active ingredients found in the plant or a natural extract.More preferably, the chemical composition of the synthetic extract hasabout 70 percent or more commonality to the chemical composition of aplant or a natural extract. Optimally, a synthetic extract has about 90percent or more commonality to the chemical composition of a plant or anatural extract.

For use in the compositions of this disclosure, the plant extract orcomponents and/or active constituents are preferably derived directlyfrom the plant. The components may be in a pure form, a semi-pure form,or unpurified form. In one embodiment, the components are in the form ofan extract obtained by organic solvent extraction.

Methods of extraction using organic solvents are known in the art andmay be used in accordance with the present invention. Briefly, oneorganic solvent extraction method involves washing and extracting theplant material using an organic solvent. Non-limiting examples oforganic solvents include methanol, ethanol, isopropanol,dichloromethane, chloroform, hexane, xylene, and petroleum ether.Well-known methods in the art may be used for organic solventextraction.

Organic solvent extraction involves collecting the raw materials fromthe plant that contain the desired constituent(s), such as seeds,needles, leaves, roots, bark, cones, stems, rhizomes, callus cells,flowers, protoplasts, organs and organ systems, and meristems. Theseplant materials are ground to small particle sizes, and then put into anextracting machine through an inlet for the raw materials by ameasurable charging machine. The plant raw material is pushed in theextracting machine by a thruster, and slowly moves the plant rawmaterial forward. Organic solvent (e.g., ethanol) may be added into themachine through a solvent inlet at the top of a waste discharge outlet.Due to the difference in gravity and equilibrium, the solvent flowstoward the raw material inlet, soaks the materials and flows out fromthe opposite side of the solvent inlet. Since the plant materials andthe solvent move in opposite directions against each other, the plantmaterials are constantly immersed in a solution that contains alow-concentration of extract. As a result of equilibrium, high yield ofplant constituent(s) may be achieved by continuously extracting theplant material against the low-concentration solution.

An extraction time adapted to remove the plant constituents is suitable,with between about 1-8 hours typical, more preferably is between about2-6 hours, and most preferably is between about 3-5 hours. Thetemperature of extraction is between about 30° C. about 90° C., betweenabout 40° C. about 70° C., and between about 50° C. about 60° C. Thecollected extract is then fine-filtered to remove debris, and may beused directly, or is concentrated, for example by distilling the solventor by other conventional processing, and the extract can also beprovided in powder form.

Similarly, aqueous-organic solvent extraction involves initiallycollecting raw materials from a plant containing the desiredalkaloid(s), such as seeds, needles, leaves, roots, bark, cones, stems,rhizomes, flowers, callus cells, protoplasts, organs and organ systems,and meristems of the plant, which are ground into small particle sizes.The ground plant material is soaked in aqueous solution that is acidicor alkaline, depending on the solubility and stability of the desiredextract under acidic or alkaline (basic) conditions. For extractionunder acidic conditions, an acid such as hydrochloric acid or sulfuricacid is added to water, e.g., at a concentration of about 3% (w/v). Forextraction under alkaline conditions, an alkali such as sodium hydroxideor sodium carbonate is added to water. The extraction time andtemperature of extraction are typically similar to that used in theorganic solvent extraction method described above.

The extract is then collected and fine-filtered to remove debris.Alkaline agents (e.g., ammonia) or acidifying agents (e.g., sulfuricacid) may be added to the extract to neutralize the solution byadjusting the pH, depending on the acidity or alkalinity of thecollected extract. The aqueous extract may be used directly,concentrated or dried. Alternatively, organic solvent may then be addedto the neutralized solution to transfer the extract actives from anaqueous phase to an organic phase. Examples of such organic solventsinclude, but are not limited to, ethanol, isopropanol, butanol,pentanol, hexanol and xylene. The extract comprising the transferredextract actives dissolved in organic solvent may be used directly, usedas a concentrate, or dried.

Extracts may also be obtained by extracting the plant material withwater, ethanol, or a mixture thereof. The hydroalcoholic solvent systemsmay comprise from about 10% by volume to about 90% by volume of ethanoland from about 10% by volume to about 90% by volume of water. Moretypically, the solvent system will comprise from about 40% by volume toabout 90% by volume of ethanol and from about 10% by volume to about 60%by volume of water.

Different plants containing different constituents may be mixed andextracted together. This process of mixed extraction may preferably beused for extracting those plants containing constituents having similarsolubility in the solvent used for extraction, such as ethanol. Themixture of extracts may be concentrated and stored in an appropriatesolvent.

The extracts can be used in admixture with physiologically acceptablesolvents, as concentrates in which a portion of the solvent or otherdiluents are removed by evaporation or other suitable physicalprocesses, or preferably the solvent can be removed to provide a driedextract. Preferably solids are removed from the extract by filtering,centrifuging, or other suitable processes. Drying may be conducted bylyphilization, spray drying, prilling, or other suitable process. Spraydrying may be conducted by combining the extraction liquid with a fillersuch as maltodextrin, followed by spray drying.

Suitable extraction processes are disclosed in PCT PublicationsWO03/079816 (describes a process for the preparation of tomato extractswith high content in lycopene), WO04/014404 (describes a process for thepreparation of an Echinacea angustifolia extract) and WO04/014958(describes extracting a polysaccharide of Echinacea angustifolia roots),all of which are herein incorporated by reference in their entirety.

Vehicle and Compositions

In accordance with this disclosure, the compositions containing theMAGP-1 actives of the invention can further comprise anti-oxidants,anti-inflammatories, sunscreens, cosmetics, including make-ups,anti-aging formulations, e.g., creams for fine lines and/or wrinkles,topicals, skin penetration enhancers, sprays, and the like. Also inaccordance with this disclosure, the plant components and additionalingredients comprising such compositions can be formulated in a varietyof product forms. Preferably, the compositions are prepared in targeteddelivery systems, e.g. creams, lotions, gels, serums, transdermalpatches, and the like, particularly for topical administration. Targeteddelivery and/or penetration enhancement may also be achieved byiontophoresis.

The present disclosure further provides the compositions comprising theplant components preferably for topical administration or for targeteddelivery without inducing significant irritation. Thus, the inventivecompositions are especially suitable for sensitive skin. Thecompositions are applied to the skin for a period of time sufficient toimprove the aesthetic appearance of skin. The compositions arepreferably applied topically once, twice, or more daily, preferably,once or twice daily. The daily application is preferably for a period ofone week, two weeks, four weeks, or more. The compositions can beformulated into liposomes or other encapsulates or delivery vehicleswhich can comprise other additives or substances, and/or which can bemodified to more specifically reach or remain at a site followingadministration.

The present disclosure also encompasses compositions comprising acosmetically or dermatologically acceptable formulation which issuitable for contact with living animal tissue, including human tissue,with virtually no adverse physiological effect to the user. Compositionsembraced by this disclosure can be provided in any cosmetically and/ordermatologically suitable form, preferably as a lotion or cream, butalso in an anhydrous or aqueous base, as well as in a sprayable liquidform. Other suitable cosmetic product forms for the compositions of thisdisclosure include, for example, an emulsion, a lip balm, a lip gloss, alotion, a mask, an ointment, a mousse, a patch, a pomade, a solution, aspray, a wax-based stick, or a towelette. In addition, the compositionscontemplated by this disclosure can include one or more compatiblecosmetically acceptable adjuvants commonly used and known by the skilledpractitioner, such as colorants, fragrances, emollients, humectants,preservatives, vitamins, chelators, thickeners, anesthetics,anti-allergenics, antifungals, antimicrobials, other anti-inflammatoryagents, antioxidants, antiseptics, depigmenting agents, film formers,insect repellents, pharmaceutical agents, photostabilizing agents,sunscreens, stabilizers, surfactants, thickeners, viscosity modifiers,and the like, as well as other botanicals such as aloe, chamomile, andthe like, and as further described below.

Cosmetically or dermatologically acceptable vehicles that can be used inthe present topical compositions include, but are not limited to, one ormore aqueous systems, glycerins, C1-4 alcohols, fatty alcohols, fattyethers, fatty esters, polyols, glycols, vegetable oils, mineral oils,liposomes, laminar lipid materials, silicone oils, water or anycombinations thereof.

In the present disclosure, the vehicle may be in the form of an aqueousphase, an oil phase, a gel, a wax-in-water emulsion, a silicone-in-wateremulsion, a water-in-silicone, an oil-in-water emulsion, or awater-in-oil emulsion. The aqueous phase is a mixture of one or morewater soluble or water dispersible ingredient, which can be liquid,semi-solid or solid at room temperature (25° C.). The vehicle comprisesor can be in the form of a suspension, dispersion or solution in wateror an aqueous-alcoholic vehicle, which may contain a thickener orgellant. A person skilled in the art can select the appropriate productform, the ingredients contained therein, as well as the method forpreparing it, on the basis of the knowledge that the skilled artisanpossesses.

The composition may include an aqueous phase which may contain water ora mixture of water and at least one hydrophilic organic solvent such asan alcohol, especially a linear or branched lower monoalcohol containingfrom 2 to 5 carbon atoms, e.g., ethanol or propanol; a polyol, e.g.,propylene glycol, sorbitol, glycerol, digylcerol, panthenol, orpolyethylene glycol, and mixtures thereof. This aqueous phase mayrepresent from about 0.5 to about 99.99 wt % by weight of thecomposition.

When the composition of the disclosure is in the form of an emulsion, itcan also optionally comprise a surfactant, preferably in an amount offrom 0.1 to 30% and in particular from about 1 to about 20 wt % byweight of the composition.

The composition can also comprise a thickening polymer such as anamphiphilic polyurethane, a polyacrylic homopolymer or copolymer, apolyester, and/or a hydrocarbon-based resin. The polymers can bedissolved or dispersed in the cosmetically acceptable vehicle andoptionally combined with a plasticizer.

The composition of the disclosure may also comprise an oil phasecontaining oil soluble or oil dispersible ingredients that are liquid atroom temperature (25° C.) and/or oily or waxy substances that are solidat room temperature, such as waxes, semisolids, gums, and mixturesthereof. This oily phase may also contain organic solvents.

Suitable oily materials that are liquid at room temperature, oftenreferred to as oils, include hydrocarbon-based oils of animal originsuch as perhydrosqualene; hydrocarbon-based plant oils such as liquidtriglycerides of fatty acids of 4 to 10 carbon atoms, for instance,heptanoic or octanoic acid triglycerides, or oils such as sunflower oil,corn oil, soybean oil, grapeseed oil, castor oil, avocado oil,caprylic/capric acid triglycerides, jojoba oil; linear or branchedhydrocarbons of mineral or synthetic origin such as liquid paraffins andderivatives thereof, petroleum jelly; synthetic esters and ethers, inparticular esters of fatty alcohols, namely, for example, isopropylmyristate, 2-ethylhexyl palmitate, 2-octyldodecyl stearate, isostearylisostearate; hydroxylated esters such as isostearyl lactate, octylhydroxystearate, octyldodecyl hydroxystearate, heptanoates, octanoatesand decanoates of fatty alcohols; polyol esters such as propylene glycoldioctanoate, neopentyl glycol diheptanoate, diethylene glycoldiisononanoate, and pentaerythritol esters; fatty alcohols containingfrom 12 to 26 carbon atoms such as octyldodecanol, 2-butyloctanol,2-hexyldecanol, 2-undecylpentadecanol, oleyl alcohol; partiallyhydrocarbon-based fluoro oils and/or fluorosilicone oils; silicone oilssuch as volatile or non-volatile, linear or cyclic polymethylsiloxanes(PDMS) that are liquid or semisolid at room temperature such ascyclomethicones and dimethicones, optionally comprising a phenyl group,for instance phenyl trimethicones, siloxanes, and mixtures thereof.These oils are usually present in an amount of 0 wt % to about 90 wt %,preferably from about 1 wt % to 80 wt % by weight of the oil phase.

The oil phase of the composition of the disclosure may also comprise oneor more cosmetically acceptable organic solvents. These solvents arepresent in an amount of 0 wt % to about 60 wt %, preferably about 1 wt %to 30 wt % by weight of the composition and can be selected from thegroup consisting of lipophilic organic solvents, amphiphilic organicsolvents and mixtures thereof. Suitable solvents which can be used inthe composition of the disclosure include acetic acid esters such asmethyl, ethyl, butyl, amyl or 2-methoxyethyl acetate; isopropyl acetate;hydrocarbons such as toluene, xylene, p-xylene, hexane or heptane;ethers containing at least 3 carbon atoms, and mixtures thereof.

The composition of the disclosure may further comprise any ingredientconventionally used in the cosmetic field. These ingredients includepreserving agents, aqueous phase thickeners (polysaccharide biopolymers,synthetic polymers) and fatty-phase thickeners, fragrances, hydrophilicand lipophilic active agents, and mixtures thereof. The amounts of thesevarious ingredients are those conventionally used in the cosmetic fieldto achieve their intended purpose, and range typically from about 0.01wt % to about 20 wt % by weight of the composition. The nature of theseingredients and their amounts must be compatible with the production ofthe compositions of the disclosure.

The composition of the disclosure may also comprise an additionalparticulate phase, typically present in an amount of 0 wt % to about 30wt % by weight of the composition, preferably from about 0.05 wt % toabout 20 wt %, and which can comprise fillers, pigments and/orpearlescent agents and/or fillers used in cosmetic compositions.Suitable inorganic pigments include titanium oxide and iron oxide;suitable fillers include talc, mica, polyethylene powder boron nitride,copolymer microspheres, and silicone resin microbeads (Tospearl fromToshiba).

The oil phase of the compositions of the disclosure may comprise one ormore waxes, gums, or mixtures thereof. The waxes includehydrocarbon-based waxes, fluoro waxes and/or silicone waxes and can beof plant, mineral, animal and/or synthetic origin. In particular, thewaxes have a melting point of greater than 25° C., preferably greaterthan 45° C. The compositions of the present disclosure may contain from0 to about 20 wt % waxes by weight of the composition. The gums aregenerally high molecular weight PDMSs or cellulose gums orpolysaccharides and the semisolid materials are generallyhydrocarbon-based compounds such as lanolins and derivatives thereof oralternatively PDMSs. The compositions of the present disclosure maycontain from 0 to about 20 wt % gums by weight of the composition,typically from about 0.1% wt to about 10 wt %.

In another preferred embodiment, the topical compositions of the presentdisclosure also include at least one of the following: a skinpenetration enhancer, a surface smoother, a skin plumper, an opticaldiffuser, a sunscreen, an exfoliation promoter, and an antioxidant.Details with respect to these and other suitable cosmetic ingredientscan be found in the “International Cosmetic Ingredient Dictionary andHandbook,” 10th Edition (2004), published by the Cosmetic, Toiletry, andFragrance Association (CTFA), at pp. 2177-2299, which is hereinincorporated by reference in its entirety.

Among the sunscreens that can be employed in the present compositionsare avobenzone, cinnamic acid derivatives (such as octylmethoxycinnamate), octyl salicylate, oxybenzone, titanium dioxide, zinc oxide,or any mixtures thereof. The sunscreen may be present from about 1 wt %to about 30 wt % of the total weight of the composition. The addition ofa sunscreen may prevent/reduce the photodegradation of the compositionwhile in the package as well as serve to protect the skin fromultraviolet radiation.

An antioxidant functions, among other things, to scavenge free radicalsfrom skin to protect the skin from environmental aggressors. Examples ofantioxidants that may be used in the present compositions include butare not limited to alpha hydroxy acids (AHA); benzoyl peroxide; betahydroxy acids; keto acids, such as pyruvic acid, 2-oxopropanoic acid,2-oxobutanoic acid, and 2-oxopentanoic acid; oxa acids as disclosed inU.S. Pat. Nos. 5,847,003 and 5,834,513 (the disclosures of which areincorporated herein by reference); salicylic acid; urea; or any mixturesthereof. The preferred antioxidant are 3,6,9-trioxaundecanedioic acid,glycolic acid, lactic acid, or any mixtures thereof; compounds havingphenolic hydroxy functions, such as ascorbic acid and itsderivatives/esters; beta-carotene; catechins; curcumin;tetrahydrocurcumin; tocopherol and its derivatives; or any mixturesthereof. Compositions of the present disclosure may have an antioxidantpreferably from about 0.001 wt % to about 10 wt %, and more preferablyfrom about 0.001 wt % to about 5 wt %, of the total weight of thecomposition.

The present composition may also have one or more of the followingactive agents, ingredients or adjuvants: anesthetics, anti-allergenics,antifungals, antiseptics, chelating agents, colorants, demulcents,emollients, emulsifiers, fragrances, humectants, lubricants,moisturizers, pH adjusters, pigment altering agents, preservatives,stabilizers, surfactants, thickeners, viscosity modifiers, vitamins, orany mixtures thereof. The amounts of these various substances are thosethat are conventionally used in the cosmetic or pharmaceutical fields,for example, they can constitute from about 0.01% to about 20% of thetotal weight of the composition.

Non-limiting examples of active agents for formulating into thecompositions of the present disclosure include those reagents having aneffect on the treatment of wrinkles and/or fine lines, in addition tothe natural plant actives as described, such as keratolytic agents,i.e., an active agent having desquamating, exfoliant, or scrubbingproperties, or an active agent which can soften the horny layer of theskin. Other examples of anti-wrinkle or anti-fine line active agentsinclude hydroxy acids and retinoids. These agents can be formulated, forexample, in amounts of from about 0.0001% to about 5% by weight relativeto the total weight of the composition.

Suitable hydroxy acids include, for example, glycolic acid, lactic acid,malic acid, tartaric acid, citric acid, 2-hydroxyalkanoic acid, mandelicacid, salicylic acid and alkyl derivatives thereof, including5-n-octanoylsalicylic acid, 5-n-dodecanoylsalicylic acid,5-n-decanoylsalicylic acid, 5-n-octylsalicylic acid,5-n-heptyloxysalicylic acid, 4-n-heptyloxysalicylic acid and2-hydroxy-3-methylbenzoic acid or alkoxy derivatives thereof, such as2-hydroxy-3-methyoxybenzoic acid.

Exemplary retinoids include, without limitation, retinoic acid (e.g.,all-trans or 13-cis) and derivatives thereof, retinol (Vitamin A) andesters thereof, such as retinol palmitate, retinol acetate and retinolpropionate, and salts thereof.

More particularly, the compositions for topical application can be inthe form of a protective care composition for the skin, preferably forthe face, the neck, the hands, the feet, or other areas of the body.Non-limiting examples include day creams or lotions, night creams orlotions, sunscreen creams, lotions, or oils, body milks, makeup (afoundation), artificial tanning compositions, depilatories, and patches.

Emulsifiers are typically present in emulsion compositions of thedisclosure in an amount of about 0.1% to about 30%, by weight andpreferably from about 0.5% to about 30% by weight relative to the totalweight

In water-based formulations, the active compounds can be formulated inthe pH range from 1 up 8, with pH=2-7 being preferred, and pH 3.5-5.5being most preferred.

In another embodiment in accordance with the present invention,compounds for stimulating the expression of MAGP-1 protein are screenedfor activity using an assay or screening method which involves culturingnormal human dermal fibroblasts in plates with DMEMD with 10% serumfollowed by incubation for 24 hours at 37° C. and 10% CO₂. After 24hours stock solutions of compounds of the present invention as testmaterials were made in appropriate solvents (e.g., DMSO). The cells arethen treated with the test materials or respective vehicle controldiluted in growth medium for 48 hours in a humidified 37° C. incubatorwith 10% CO₂. After incubation, growth medium is typically removed and aTRIzol reagent is added to the plates. The resulting cell lysates arecollected and placed in a freezer until RNA isolation and analysis. RNAisolation can be performed using any method known to those of skill inthe art. After isolation, concentration of total RNA is determined byNanodrop (Thermo Scientific, DE) or any other known method. The isolatedRNA then undergoes Reverse Transcription (RT) and quantitativePolymerase Chain Reaction (qPCR) to amplify the RNA, followed bymeasurement and calculation according to known methods to determine thelevel of MAGP-1 stimulation activity of each compound.

A further embodiment of the invention is to prepare compositionscontaining a MAGP-1 active identified in accordance with theabove-disclosed screening method.

The following examples are meant to demonstrate certain aspects of theinvention in a non-limiting fashion.

EXAMPLES Example 1 Preparation of Extracts

Ixora chinensis—An extract is obtained by extracting the dry choppedplant of Ixora chinensis Lamk. plant using an ethanol extractionfollowed by a further extraction with hexane. Briefly, the chopped plantof Ixora chinensis is first manually ground into small particlesresulting in a powder of about 250 grams. The ground powder is thenextracted with 50% ethanol. After filtering and vacuum evaporation, thetotal concentrated extract is diluted with water, centrifuged andfiltered. The liquid is then thrice extracted with hexane, the hexaneupper layer being discarded and the aqueous layer being lyophilizedresulting in an extract of about 90 grams.

Clerodendron linleyi—An extract is obtained by extracting leaves of theClerodendron linleyi plant using an ethanol extraction scheme. Theleaves of Clerodendron linleyi are first manually ground into smallparticles resulting in a powder of about 250 grams per flask (2 flasks).The ground powder is then extracted with 80 vol % ethanol (2×2,000 mlper flask). After filtering and vacuum evaporation, the totalconcentrated extract is lyophilized resulting in an ethanolic extract of30.77 grams.

Antidesma bunius—An extract is obtained from 400 grams of ground plantleaves extracted with 4450 ml of a 1:1 ethanol-water solution by volumein three portions. The extract is filtered, concentrated under vacuum toremove ethanol, diluted with water to 1300 ml, and then centrifuged. Theresulting supernatant liquid is washed with 500 ml of dichloromethane,decolorized with 10 g of charcoal, filtered, then concentrated to yield57.7 g of extract, which is dried to a powder.

Operculina turphetum—An extract is obtained by extracting the aerialportions of the Operculina turphetum plant using an ethanol extractionscheme. The leaves of Operculina turphetum are first manually groundinto small particles resulting in a powder of about 250 grams. Theground powder is then extracted with a water-ethanol mixture (50-50).After filtering and vacuum evaporation, the aqueous solution is washedwith methylene chloride, concentrated under vacuum and dried.

Tiliacora triandra—An extract is obtained by extracting the vine of theTiliacora triandra plant using an ethanol extraction scheme. Briefly,the vines of Tiliacora triandra Diels are first manually ground intosmall particles resulting in a powder of about 250 grams per flask (2flasks). The ground powder is then extracted with 80% ethanol (2×2,000ml per flask). After filtering and vacuum evaporation, the totalconcentrated extract ias lyophilized resulting in an ethanolic extractof 50 grams.

Example 2 MAGP-1 Expression in Dermis Materials and Methods

Skin biopsies were obtained from both photo-exposed and photo-protectedareas of 15 younger subjects (age 18-25) and 15 older subjects (age40-65). Tissues were fixed in formalin and subject toimmunohistochemistry study. Anti-MAGP-1 and Alkalinephosphatase-conjugated secondary antibody were applied accordingly tovisualize protein localization in skin. Images were taken by ZeissAxionskop2 microimaging system.

Results

MAGP-1 protein is structurally degraded with photoaging (FIG. 1). Thearrow in FIG. 1 indicates the absence of MAGP-1 fibrils in papillarydermis. The star indicates structural degradation of MAGP-1 in dermis.

Example 3 MAGP-1 Expression Around Pores and Hair Follicles

Using similar materials and methods as set out in Example 2, the resultshere demonstrated that MAGP-1 protein level is significantly reducedaround hair follicle/pores with photoaging (FIG. 2). The arrows in FIG.2 indicate high levels of MAGP-1 fibril structure around hairfollicle/pore from younger subjects; and arrowheads indicate the absenceof MAGP-1 around hair follicle/pore in photoexposed skin from oldersubjects.

Example 4 Screening Method Cell Treatment

Normal human dermal fibroblasts (Cascade Biologics) were cultured in 100mm plates with 10 ml of DMEM with 10% serum and incubated for 24 hoursat 37° C. and 10% CO₂. After 24 hours, stock solutions of testcompounds/plant extracts were made in appropriate solvents (e.g., DMSO).Cells were treated with test material or respective vehicle controldiluted in growth medium for 48 hours in a humidified 37° C. incubatorwith 10% CO₂. After incubation, growth medium from each plate wasremoved and 800 ul of TRIzol Reagent (Invitrogen) was added to theplates. Cell lysates were collected and placed in −80° C. freezer untilRNA isolation and analysis.

RNA Isolation

RNA was isolated by following the manufacturer's procedures. Briefly,the cell lysates were brought to room temperature. 0.2 ml of chloroformper 1 ml of TRIzol® Reagent was added. The tubes were shaken vigorouslyfor 15 seconds and incubated at ambient temperature for 2 to 3 minutes.The samples were centrifuged at 12,000×g for 15 minutes at 2 to 8° C.Following centrifugation; the mixture separated into multiple phases,and RNA remains exclusively in the upper aqueous phase.

The aqueous phase was then transferred to a fresh tube. The RNA wasprecipitated from the aqueous phase by mixing with 0.5 ml of isopropylalcohol per 1 ml of TRIzol® Reagent was used for the initialhomogenization. Samples were incubated at 15 to 30° C. for 10 minutesand centrifuged at 12,000×g for 10 minutes at 2 to 8° C. The supernatantwas removed, and the RNA pellet was washed once with 75% ethanol byadding at least 1 ml of 75% ethanol per 1 ml of TRIzol® Reagent used forthe cell lysate. The sample was mixed by vortexing and centrifuged at7,500×g for 5 minutes at 2 to 8° C. The RNA pellet was briefly airdried. RNA was dissolved in RNase-free water, and the concentration oftotal RNA was determined by using Nanodrop (Thermo Scientific, DE).

Reverse Transcriptase and qPCR

Reverse Transcription (RT) reactions were conducted in a total volume of20 μl. The RT mixture was prepared to contain 2 μl 10× TaqMan RT buffer,4.4 μl 25 mM MgCl2, 4.0 μl dNTP mix, 1.0 μl Random Hexamer, 0.4 μl RNaseInhibitor, 0.5 μl MultiScribe Reverse Transcriptase (50 U/ml), 2.0 μg ofRNA, and RNase-free water to make up the final volume of 20 μl. Thereaction was incubated at 25° C. for 10 min, 48° C. for 30 min, then 95°C. for 5 min using Stratagene Mx 3005P QPCR machine. After the RT step,the reactions were stored at −20° C. for qPCR analysis.

Quantitative Polymerase Chain Reaction (qPCR) was conducted in a totalvolume of 20 μl. Applied Biosystems (AB) Universal PCR master mix wasused. The mixture was prepared to contain 10 μl of Taqman Universal PCRmix, 1 μl of primer and probe mix, 2 μl of RT product, and 7 μl ofdeionized water. Both MAGP-1 and GAPDH primers and probes were purchasedfrom Applied Biosystems, and ID for MAGP-1 is Hs00250064_ml, and Partnumber for human GAPDH is 4352934E.

The temperature profiles for QPCR were 50° C. for 2 min, and 95° C. for10 min for 1 cycle, then at 95° C. for 15 sec, and 60° C. for 1 min for40 cycles. Values for MAGP-1 mRNA were collected along with those forthe reference genes GAPDH. Values are normalized to GAPDH to determinechanges in MAGP-1 mRNA after treatment. Percent increase in mRNA forMAGP-1 was calculated by comparing the test results to the vehiclecontrol.

Table 1 lists ingredients that were determined to be active atstimulating expression of MAGP-1.

TABLE 1 MAGP-1 Screening Data Activity Level Product/Ingredient NameConcentration (see Key) KTFK 0.001% ++++ Amorphophallus campanulatus0.1% ++++ Sesbania grandiflora 0.01% + Thiodipropionic acid (TDPA)0.001% ++ Sapindus rarak 0.001% ++ Compound of structure I 0.01% ++++Antidesma bunius 0.10% +++ Tiliacora triandra 0.01% +++ Compound ofstructure II 0.01% ++++ Thunbergia laurifolia 0.01% + Operculinaturpethum 0.00% ++ Ixora chinensis 0.01% +++ Clerodendron lindleyi0.001% + Key to activity 0: 0-20% +: 21-40% ++: 41-60% +++: 61-80% ++++:>80%

All percentages are by weight, based on the total weight of thecomposition, unless otherwise indicated.

All references including patent applications and publications citedherein are incorporated herein by reference in their entirety and forall purposes to the same extent as if each individual publication orpatent or patent application was specifically and individually indicatedto be incorporated by reference in its entirety for all purposes. Manymodifications and variations of this invention can be made withoutdeparting from its spirit and scope, as will be apparent to thoseskilled in the art. The specific embodiments described herein areoffered by way of example only, and the invention is to be limited onlyby the terms of the appended claims, along with the full scope ofequivalents to which such claims are entitled.

What is claimed is:
 1. A composition comprising a MAGP-1 stimulatingsubstance in a cosmetically acceptable vehicle, wherein the activity ofsaid MAGP-1 stimulating substance has been identified by a MAGP-1screening assay.
 2. A composition for improving the aesthetic appearanceof human skin comprising: (i) a MAGP-1 active agent selected from thegroup consisting of: an extract of Antidesma bunius, an extract ofOperculina turpethum, an extract of Ixora chinensis, an extract ofClerodendron lindleyi, a compound of formula II:

and combinations thereof, and (ii) a cosmetically acceptable vehicle. 3.The composition according to claim 2, wherein the composition is in theform of an aerosol spray, pump spray, cream, emulsion, solid, liquiddispersion, foam, gel, lotion, mousse, ointment, powder, patch, pomade,serum, stick or towlette.
 4. The composition according to claim 2,wherein the MAGP-1 active agent is present in an amount of from about0.01 wt % to about 10 wt % of the total weight of the composition. 5.The composition according to claim 2, comprising Antidesma bunius. 6.The composition according to claim 2, comprising Operculina turpethum.7. The composition according to claim 2, comprising Ixora chinensis 8.The composition according to claim 2, comprising Clerodendron linleyi.9. The composition according to claim 2, comprising a compound havingthe formula:


10. The composition according to claim 2, further comprising a secondMAGP-1 active agent selected from the group consisting of extracts ofSesbania grandiflora, extracts of Amorphophallus campanulatus, extractsof Sapindus rarak, extracts of Tiliacora triandra, extracts ofThunbergia laurifolia, KFTK, thiodipropionic acid and its C₁-C₄ mono-and dialkyl esters, and combinations thereof.
 11. A method for improvingthe condition or aesthetic appearance of human skin comprising topicallyapplying to skin in need thereof an effective amount of a composition ofclaim 2, wherein said improvement is selected from the group consistingof: (a) treatment, reduction, and/or prevention of fine lines orwrinkles, (b) reduction of skin pore size, (c) improvement in skinthickness, plumpness, and/or firmness; (d) improvement in skinsuppleness and/or softness; (e) improvement in skin tone, radiance,and/or clarity; (f) improvement in procollagen and/or collagenproduction; (g) improvement in maintenance and remodeling of elastin;(h) improvement in skin texture and/or promotion of retexturization; (i)improvement in skin barrier repair and/or function; (j) improvement inappearance of skin contours; (k) restoration of skin luster and/orbrightness; (l) replenishment of essential nutrients and/or constituentsin the skin; (m) improvement of skin appearance decreased by menopause;(n) improvement in skin moisturization; (o) increase in skin elasticityand/or resiliency; (p) treatment, reduction, and/or prevention of skinsagging; or (r) reduction of pigment spots; or any combination thereof.12. The method according to claim 11, wherein said improvement is thetreatment, reduction, and/or prevention of fine lines or wrinkles. 13.The method according to claim 11, wherein said improvement is anincrease in skin elasticity and/or resiliency.
 14. The method accordingto claim 11, wherein said improvement is the treatment, reduction,and/or prevention of skin sagging.
 15. The method according to claim 11,wherein the composition is topically applied at least once daily for atleast one week.
 16. A method for improving the condition or aestheticappearance of human skin comprising topically applying to skin in needthereof an effective amount of a composition of claim 2, wherein saidimprovement is selected from the group consisting of: (a) treatment,reduction, and/or prevention of fine lines or wrinkles, (b) reduction ofskin pore size, (c) improvement in skin thickness, plumpness, and/orfirmness; (d) improvement in skin suppleness and/or softness; (e)improvement in skin tone, radiance, and/or clarity; (f) improvement inprocollagen and/or collagen production; (g) improvement in maintenanceand remodeling of elastin; (h) improvement in skin texture and/orpromotion of retexturization; (i) improvement in skin barrier repairand/or function; (j) improvement in appearance of skin contours; (k)restoration of skin luster and/or brightness; (l) replenishment ofessential nutrients and/or constituents in the skin; (m) improvement ofskin appearance decreased by menopause; (n) improvement in skinmoisturization; (o) increase in skin elasticity and/or resiliency; (p)treatment, reduction, and/or prevention of skin sagging; or (r)reduction of pigment spots; or any combination thereof.
 17. The methodaccording to claim 16, wherein said improvement is the treatment,reduction, and/or prevention of fine lines or wrinkles.
 18. The methodaccording to claim 16, wherein said improvement is the treatment,reduction, and/or prevention of skin sagging.
 19. The method accordingto claim 16, wherein the composition is topically applied at least oncedaily for at least one week.